The finding of enzymes capable of catalyzing different reactions with high efficiency, stability and specificity can have origin from extremophile organisms, which are those that dwell in environment known as extreme. The enzymes of these organisms can have physical and chemical characteristics giving them generally stability to extreme conditions in which the organism that generates it lives.
The extremophile organisms as well as thermoacidophiles organisms can generate active enzymes in most cases, with low pH's and high temperatures. The acidophile organisms are known by the production of stable proteins such as proteases of Sulfolobus acidocaldarius (Fusek et. al. 1990), a-amylase of Bacilus acidicola (Serour & Satyanarayana, 2011), glucoamylases of Picrophilus torridus (Serour & Antranikian, 2002) and esterases of Ferroplasma acidiphilum (Golyshina et. al. 2006).
Hydrolytic enzymes esterases and lipases type have been described of thermoacidophiles mainly of the archaeans domain (Kim & Lee, 2004). Within the domain bacteria it is only known an esterase described from a thermoacidophile organism, Alicyclobacillus acidocaldarius, (Manco, et. al. 1998).
The importance of the lipolytic enzymes lies in their versatility when intervening in different catalytic processes, both of synthesis or hydrolysis of ester links, which can be applicable to different biotechnological processes. Among the properties of these enzymes are their high chemo, regio and enantio specificity. Biotechnologically, they can be useful in the synthesis of polymers, biodiesel, agrochemical products, flavoring compounds, hydrolysis of intermediate synthesis, or building blocks and besides in the resolution of racemic mixtures. (Hassanm, et al., 2006, Joseph et al. 2008
Due to its high degree of enantioselectivity, lipolytic enzymes are used in the resolution of racemic mixtures of compounds such as R/S-ibuprofen that belong to the family of the NSAIDs. Despite all the benefits, most of these medicaments have been commercialized as racemic mixtures mainly due to the high costs of separation of the enantiomers. The obtainment of enantiomers S-ibuprofen o S-naproxen show better benefits than the racemic mixture, for this reason it is important to find biological mechanisms of obtainment of simple enantiomers and this justifies the interest developed in recent decades for the obtainment of S (+)-2-pure aryl propionic acids. Now the transformation of the racemic mixture to obtain the S pure enantiomer is carried out through hydrolases, specifically lipolytic enzymes.
The Acidophilus thermo organism USBA-GBX-499 from which the basal esterase was obtained was isolated from an acid thermal spring from the National Natural Park of Los Nevados. This organism grows within a temperature range between 40° C. and 70° C. and a pH from 2.0 to 5.0 and degrades different substrata such as tributyrin, tricaprylin, ethyl oleate and others. From the extracellular fraction of the strain USBA-GBX-499 it was detected an enzymatic lipolytic activity from 0.01 U/mg. This low value indicated the presence of a thermostable lipolytic enzyme. Therefore, it was made the identification, clonation, expression and biochemical characterization of the thermo alkaline lipolytic enzyme called 499EST, which is modified in an intermediate stage of the purification process of the enzyme, thus obtaining the modified esterase enzyme which is the object of this invention.
Within the characterization of 499EST, it was identified an evaluation of enantioselectivity, its preference for the enantiomers (S)-ibuprofen and (S)-naproxen, which are enantiomers of high importance in the pharmaceutical industry.